Novel SCN5A mutation in amiodarone-responsive multifocal ventricular ectopy-associated cardiomyopathy

Abstract

Background: Mutations in SCN5A, which encodes the cardiac sodium channel NaV1.5, typically cause ventricular arrhythmia or conduction slowing. Recently, SCN5A mutations have been associated with heart failure combined with variable atrial and ventricular arrhythmia.

Objective: The purpose of this study was to determine the clinical, genetic, and functional features of an amiodarone-responsive multifocal ventricular ectopy-related cardiomyopathy associated with a novel mutation in a NaV1.5 voltage sensor domain.

Methods: A novel, de novo SCN5A mutation (NaV1.5-R225P) was identified in a boy with prenatal arrhythmia and impaired cardiac contractility followed by postnatal multifocal ventricular ectopy suppressible by amiodarone. We investigated the functional consequences of NaV1.5-R225P expressed heterologously in tsA201 cells.

Results: Mutant channels exhibited significant abnormalities in both activation and inactivation leading to large, hyperpolarized window and ramp currents that predict aberrant sodium influx at potentials near the cardiomyocyte resting membrane potential. Mutant channels also exhibited significantly increased persistent (late) sodium current. This profile of channel dysfunction shares features with other SCN5A voltage sensor mutations associated with cardiomyopathy and overlapped that of congenital long QT syndrome. Amiodarone stabilized fast inactivation, suppressed persistent sodium current, and caused frequency-dependent inhibition of channel availability.

Conclusion: We determined the functional consequences and pharmacologic responses of a novel SCN5A mutation associated with an arrhythmia-associated cardiomyopathy. Comparisons with other cardiomyopathy-associated NaV1.5 voltage sensor mutations revealed a pattern of abnormal voltage dependence of activation as a shared biophysical mechanism of the syndrome.

Keywords: Amiodarone; Cardiomyopathy; Electrophysiology; SCN5A mutation.

Figure 1. Electrocardiograms from the proband

(A) Representative lead II recording obtained at age 3 days illustrating wandering atrial rhythm with frequent multifocal premature ventricular beats. (B) Representative lead II recording obtained at age 7 weeks illustrating an episode of supraventricular tachycardia with variable 2:1 and 3:1 AV block. (C) Representative lead II recording obtained at age 6.5 months two weeks off amiodarone illustrating sinus rhythm with a prolonged QTc (480 ms). (D) Representative lead II recording obtained at age 9 months while the proband was off antiarrhythmic drugs illustrating multifocal ventricular ectopy and two 3–4 beat runs of nonsustained ventricular tachycardia. (E) Representative lead II recording obtained at age 20 months while treated with amiodarone and propranolol illustrating sinus rhythm. More complete ECG recordings corresponding to each of these events are provided as Supplemental Fig. S2.

Figure 2. Biophysical properties of R225P

Biophysical properties of Na_V1.5-R225P. (A) Representative traces of WT (top) and R225P (bottom) sodium channels. (B) Current-density/voltage plots of WT and R225P. (C) Voltage dependence of activation for WT and R225P from −80 to +20 mV. (D) Representative traces of WT and R225P illustrating altered activation and inactivation kinetics. (E) Voltage-dependence of inactivation time constants (open symbols represent fast component; closed symbols represent slow component) for WT and R225P. (F) Representative TTX-subtracted whole cell current for WT and R225P. Persistent current was measured over the final 10 ms of a 200 ms pulse to −20 mV and normalized to peak current. Inset shows persistent current over the final 50 ms. All data are represented as mean ± S.E.M for n=11–18 cells.

Figure 3. Window- and Ramp-currents

Window and ramp-currents of Na_V1.5-R225P reveal aberrant I_Na at hyperpolarized potentials. (A) Overlay of Boltzmann-fitted G/V and channel availability curves of WT and R225P emphasizing window-currents. (B) Normalized TTX-subtracted average ramp-currents (0.32 mV/ms) of WT and R225P measured as a percentage of peak I_Na (−20mV). All data are represented as mean ± S.E.M for n=7–15 cells.

Figure 4. Frequency-dependent channel rundown and slow inactivation

Na_V1.5-R225P stabilizes the slow-inactivated state. (A) Frequency-dependent channel rundown of WT (black) and R225P (red) observed for activating pulse durations of 5, 100, and 300 ms. Curves were fit with exponential decay functions. (B) Onset of slow inactivation for WT and R225P. (C) Recovery from slow inactivation. WT (black line) and R225P (red line) data fit with double-exponential curves. (D) Voltage-dependence of slow-inactivation of WT and R225P. Curves are the result of data fittings with the Boltzmann function. All data are represented as mean ± S.E.M for n=7–15 cells.

Figure 5. Normalized ramp-currents

Comparison of normalized ramp-currents. Normalized TTX-subtracted averaged ramp-currents from R225P and other Na_V1.5 mutations associated with multifocal ventricular ectopy with impaired cardiac contractility (R222Q), sporadic dilated cardiomyopathy (R814W), and congenital long-QT syndrome (delKPQ). All data are represented as mean for n=7–15 cells.

Figure 6. Biophysical properties of R225P in the presence of Amiodarone (3 μM)

Effects of amiodarone on Na_V1.5-R225P. (A) Activation and channel availability properties of WT and R225P in the presence of amiodarone (open symbols, dashed lines) or DMSO (filled circles, solid lines). All data were fit with a Boltzmann function. (B) TTX-subtracted averaged ramp-currents (0.32 mV/ms) of WT and R225P normalized to cell capacitance in the presence of amiodarone (dashed lines) or DMSO (solid lines). (C) Recovery from fast inactivation of WT and R225P in the presence of amiodarone (open symbols, dashed lines) or DMSO (filled circles, solid lines). The data were fit with a double exponential curve. (D) Pulse trains (2Hz; −20mV) with an activating pulse of 100ms of WT and R225P channels in the presence of amiodarone (open symbols, dashed lines) or DMSO (filled circles, solid lines). All data are represented as mean ± S.E.M for n=5–12 cells.